Use of semenogelin in the diagnosis, prognosis and treatment of cancer

ABSTRACT

Method of diagnosing cancer in a male mammal comprising obtaining and assaying a test sample for an increased level of semenogelin method of diagnostic cancer in a female mammal comprising obtaining and assaying a test sample for the presence of semenogelin methods of prognosticating and assessing the effectiveness of treatment of a cancer in a mammal comprising measuring the level of semenogelin in a test sample method of inducing an immune response to a cancer in a mammal comprising administering to the mammal a composition comprising (a) an immune-response inducing effective amount of (i) semenogelin or (ii) antibody thereto or (b) a recombinant vector encoding and expressing an immune-response inducing effective amount of (i) or (ii); and composition comprising a carrier and (a) an immune-response inducing effective amount of (i) a polypeptide of any of SEQ ID NOS:1-27 or (ii) antibody thereto or (b) a recombinant vector encoding and expressing an immune-response inducing effective amount of (i) or (ii).

FIELD OF THE INVENTION

[0001] This invention pertains to the use of semenogelin in thediagnosis, prognosis and treatment of cancer, particularly small-celllung cancer.

BACKGROUND OF THE INVENTION

[0002] The American Cancer Society estimates the lifetime risk that anindividual will develop cancer is 1 in 2 for men and 1 in 3 for women.The development of cancer, while still not completely understood, can beenhanced as a result of a variety of risk factors. For example, exposureto environmental factors (e.g., tobacco smoke) might triggermodifications in certain genes, thereby initiating cancer development.Alternatively, these genetic modifications may not require an exposureto environmental factors to become abnormal. Indeed, certain mutations(e.g., deletions, substitutions, etc.) can be inherited from generationto generation, thereby imparting an individual with a geneticpredisposition to develop cancer.

[0003] Currently, the survival rates for many cancers are on the rise.One reason for this success is improvement in the detection of cancer ata stage at which treatment can be effective. Indeed, it has been notedthat one of the most effective means to survive cancer is to detect itspresence as early as possible. According to the American Cancer Society,the relative survival rate for many cancers would increase by about 15%if individuals participated in regular cancer screenings. Therefore, itis becoming increasingly useful to develop novel diagnostic tools todetect the cancer either before it develops or at an as early stage ofdevelopment as possible.

[0004] One popular way of detecting cancer early is to analyze thegenetic makeup of an individual to detect the presence or expressionlevels of a marker gene(s) related to the cancer. For example, there arevarious diagnostic methods that analyze a certain gene or a pattern ofgenes to detect cancers of the breast, tongue, mouth, colon, rectum,cervix, prostate, testis, and skin. Recently, measuring the level ofexpression of semenogelin has been found to be useful in the detectionof prostate cancer.

[0005] Semenogelin is known to be the predominant protein in humansemen. Normally, it is synthesized by the secretory epithelium of theseminal vesicles as a 461 amino acid precursor protein. Followingcleavage of a predominantly hydrophobic signal peptide, the secretedprotein contains 439 amino acid residues. Semenogelins I and II(hereinafter referred to as “SgI” and “SgII,” respectively) are twoseparate gene products resulting from the expression of semenogelin andare normally responsible for the gel formation in semen. SgI is asingle-chain, non-glycosylated protein of 439 amino acids, whereas SgIIcontains 559 amino acids (Lilja et al., Proc. Natl. Acad. Sci. USA. 89(10): 4559-4563 1992). Degradation of SgI and SgII is due to theproteolytic action of prostate-specific antigen (PSA), which also hasbeen shown to be helpful in diagnosing prostate cancer.

[0006] While semenogelin has been detected in human semen, histologicalanalyses have failed to detect semenogelin expression in any normalhuman tissue other than seminal vesicle epithelium (Herr et al., BiolReprod. 40: 333-342 (1989); Evans et al., Anat Rec. 214: 372-377 (1986);and Bjartell et al., J Androl. 17: 17-26 (1996)). Since semenogelinexpression occurs normally in seminal vesicle epithelium, it has beenshown that a decrease in expression of semenogelin in seminal vesicleepithelium is indicative of prostate cancer (see, e.g., U.S. Pat. No.5,972,615).

[0007] A need remains for additional ways to diagnose, prognosticate,and treat cancer. The invention provides such methods. These and otheradvantages of the invention, as well as additional inventive features,will be apparent from the description of the invention provided herein.

BRIEF SUMMARY OF THE INVENTION

[0008] The invention provides a method of diagnosing cancer in a malemammal, wherein the cancer is other than prostate cancer, which methodcomprises obtaining a test sample from the male mammal and subsequentlyassaying the test sample for an increased level of semenogelin, whereinthe increased level of semenogelin in the test sample is diagnostic forthe cancer. The invention also provides a method of diagnosing cancer ina female mammal, which method comprises obtaining a test sample from thefemale mammal and subsequently assaying the test sample for the presenceof semenogelin, wherein the presence of semenogelin in the test sampleis diagnostic for the cancer. Also provided by the invention is a methodof prognosticating a cancer in a mammal, wherein the cancer is otherthan prostate cancer and semenogelin is a marker for the cancer, whichmethod comprises measuring the level of semenogelin in a test sampleobtained from the mammal, wherein the level of semenogelin in the testsample is indicative of the prognosis of the cancer in the mammal.Further provided is a method of assessing the effectiveness of treatmentof a cancer in a mammal, wherein the cancer is other than prostatecancer and semenogelin is a marker for the cancer, which methodcomprises measuring the level of semenogelin in a test sample obtainedfrom the mammal, wherein the level of semenogelin in the test sample isindicative of the effectiveness of the treatment of the cancer in themammal. Still further provided is a method of inducing an immuneresponse to a cancer in a mammal, wherein the cancer is other thanprostate cancer and semenogelin is a marker for the cancer, which methodcomprises administering to the mammal a composition comprising (a) animmune-response inducing effective amount of (i) a semenogelin proteinor polypeptide fragment thereof or (ii) an antibody or antigenicallyreactive fragment thereof that is specific for a semenogelin protein orpolypeptide fragment thereof or (b) a recombinant vector encoding andexpressing an immune-response inducing effective amount of (i) or (ii),whereupon an immune response to the cancer is induced. The inventionfurther provides a composition comprising a pharmaceutically acceptablecarrier and (a) an immune-response inducing effective amount of (i) apolypeptide of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ IDNO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ IDNO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ IDNO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ IDNO:25, SEQ ID NO:26, or SEQ ID NO:27 or (ii) an antibody orantigenically reactive fragment thereof that is specific for apolypeptide of SEQ ID NO: 1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ IDNO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ IDNO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ IDNO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ IDNO:25, SEQ ID NO:26, or SEQ ID NO:27 or (b) a recombinant vectorencoding and expressing an immune-response inducing effective amount of(i) or (ii).

DETAILED DESCRIPTION OF THE INVENTION

[0009] The invention provides a method of diagnosing cancer in a malemammal, wherein the cancer is other than prostate cancer. The methodcomprises: (a) obtaining a test sample from the male mammal, and (b)assaying the test sample for an increased level of semenogelin, whereinthe increased level of semenogelin in the test sample is diagnostic forthe cancer. The test sample can be assayed for an increased level ofsemenogelin in (b) by comparing the level of semenogelin in the testsample to the level of semenogelin in a control sample obtained from oneor more cancer-free male mammals of the same species, wherein anincrease in the level of semenogelin in the test sample as compared tothe control sample is diagnostic for the cancer. Alternatively, thelevel of semenogelin in the test sample can be compared to an alreadydetermined range of semenogelin for cancer-free male mammals of the samespecies.

[0010] In addition, the invention provides a method of diagnosing cancerin a female mammal. The method comprises: (a) obtaining a test samplefrom the female mammal, and (b) assaying the test sample for thepresence of semenogelin, wherein the presence of semenogelin in the testsample is diagnostic for the cancer.

[0011] For purposes of the invention, when a female mammal is beingdiagnosed, the presence of any cancer can be assayed for. A male mammal,however, can be assayed for any cancer other than prostate cancer.Preferably, the cancer is of epithelial origin, such as lung cancer,papillary renal cell carcinoma, colon cancer, and melanoma. Mostpreferably, the cancer is small-cell lung cancer (hereinafter referredto as “SCLC”).

[0012] The test sample used in conjunction with the invention can be anyof those typically used in the art. For example, the test sample can betissue. Typically, the tissue is metastatic (e.g., cancerous) and isobtained by means of a biopsy. Such tissue can include bone marrow,lymph nodes, skin, and any organ that may develop cancerous cells.Preferably, however, the test sample is taken from a source in whichsecreted proteins will be most prevalent. Accordingly, the test sampleis preferably serum, wherein the serum is obtained from methods known inthe art, such as a blood sample.

[0013] A number of assays are contemplated for use in the presentinventive methods of diagnosing cancer. A number of these assays aredescribed in Sambrook et al., Molecular Cloning: A Laboratory Manual,2^(nd) Ed., Cold Spring Harbor Press, Cold Spring Harbor, N.Y., 1989.Microarrays, such as those described in U.S. Pat. Nos. 6,197,506 and6,040,138, also can be used to detect and quantify semenogelin. It willbe understood that the type of assay used will depend on whether DNA,RNA or a protein (or a polypeptide thereof) is being assayed.

[0014] As used herein, the term “increased level” can be defined asdetecting semenogelin in a male mammal at a level above that which isconsidered normal. For example, the level of semenogelin in a testsample is increased when the copy number of the gene encoding thesemenogelin is greater than 1, the mRNA encoding semenogelin is about0.001-1%, or semenogelin (or a polypeptide thereof) is detected in anamount of about 1-10,000 ng/ml.

[0015] When a nucleic acid (i.e., DNA or RNA) is assayed, various assayscan be used to measure the presence and/or level of nucleic acidpresent. For example, when only the detection of semenogelin isnecessary to diagnose effectively the cancer, such as in diagnosing afemale mammal, assays including PCR and microarray analysis can be used.In certain embodiments, such as in diagnosing a male mammal, it will benecessary to detect the quantity of semenogelin present. In theseembodiments, it will be advantageous to use various hybridizationtechniques known in the art that can effectively measure the level ofsemenogelin in a test sample. When the semenogelin comprises DNA, suchhybridization techniques can include, for example, Southernhybridization (i.e., a Southern blot), in situ hybridization andmicroarray analysis. Similarly, when the semenogelin comprises RNA,Northern hybridization (i.e., a Northern blot), in situ hybridizationand microarray analysis are contemplated.

[0016] It will be understood that, in such assays, a nucleic acidsequence that specifically binds to or associates with a nucleic acidencoding semenogelin, whether DNA or RNA, can be attached to a label fordetermining hybridization. A wide variety of appropriate labels areknown in the art, including fluorescent, radioactive, and enzymaticlabels as well as ligands, such as avidin/biotin, which are capable ofbeing detected. Preferably, a fluorescent label or an enzyme tag, suchas urease, alkaline phosphatase or peroxidase, is used instead of aradioactive or other environmentally undesirable label. In the case ofenzyme tags, colorimetric indicator substrates are known which can beemployed to provide a detection means visible to the human eye orspectrophotometrically to identify specific hybridization withcomplementary semenogelin nucleic acid-containing samples.

[0017] When a nucleic acid encoding the semenogelin is amplified in thecontext of a diagnostic application, the nucleic acid used as a templatefor amplification is isolated from cells contained in the test sample,according to standard methodologies. (Sambrook et al., MolecularCloning: A Laboratory Manual, 2^(nd) Ed., Cold Spring harbor Press, ColdSpring harbor, N.Y., 1989). The nucleic acid can be genomic DNA orfractionated or whole cell RNA. Where RNA is used, it can be desirableto convert the RNA to cDNA.

[0018] In a typical amplification procedure, pairs of primers thatselectively hybridize to nucleic acids corresponding to semenogelin arecontacted with the nucleic acid under conditions that permit selectivehybridization. Once hybridized, the nucleic acid-primer complex iscontacted with one or more enzymes that facilitate template-dependentnucleic acid synthesis. Multiple rounds of amplification, also referredto as “cycles,” are conducted until a sufficient amount of amplificationproduct is produced.

[0019] Various template-dependent processes are available to amplify thesemenogelin present in a given test sample. As with the various assays,a number of these processes are described in Sambrook et al. (1989),supra. One of the best-known amplification methods is the polymerasechain reaction (PCR). Similarly, a reverse transcriptase PCR (RT-PCR)can be used when it is desired to convert mRNA into cDNA. Alternativemethods for reverse transcription utilize thermostable DNA polymerasesand are described in WO 90/07641, for example.

[0020] Other methods for amplification include the ligase chain reaction(LCR), which is disclosed in U.S. Pat. No. 4,883,750; isothermalamplification, in which restriction endonucleases and ligases are usedto achieve the amplification of target molecules that contain nucleotide5′-[alpha-thio]-triphosphates in one strand (Walker et al., Proc. Natl.Acad. Sci. USA 89: 392-396 (1992)); strand displacement amplification(SDA), which involves multiple rounds of strand displacement andsynthesis, i.e., nick translation; and repair chain reaction (RCR),which involves annealing several probes throughout a region targeted foramplification, followed by a repair reaction in which only two of thefour bases are present. The other two bases can be added as biotinylatedderivatives for easy detection. Target-specific sequences also can bedetected using a cyclic probe reaction (CPR). In CPR, a probe having 3′and 5′ sequences of non-specific DNA and a middle sequence of specificRNA is hybridized to DNA, which is present in a sample. Uponhybridization, the reaction is treated with RNase H, and the products ofthe probe are identified as distinctive products, which are releasedafter digestion. The original template is annealed to another cyclingprobe and the reaction is repeated. A number of other amplificationprocesses are contemplated; however, the invention is not limited as towhich method is used.

[0021] Following amplification of the semenogelin, it can be desirableto separate the amplification product from the template and the excessprimer for the purpose of determining whether specific amplification hasoccurred. In one embodiment, amplification products are separated byagarose, agarose-acrylamide or polyacrylamide gel electrophoresis usingstandard methods. See Sambrook et al. (1989), supra.

[0022] Alternatively, chromatographic techniques can be employed toeffect separation. There are many kinds of chromatography which can beused in the context of the present inventive methods e.g., adsorption,partition, ion-exchange and molecular sieve, and many specializedtechniques for using them including column, paper, thin-layer and gaschromatography (Freifelder, Physical Biochemistry Applications toBiochemistry and Molecular Biology, 2^(nd) Ed., Wm. Freeman and Co., NewYork, N.Y. (1982)).

[0023] Amplification products must be visualized in order to confirmamplification of the semenogelin sequence. One typical visualizationmethod involves staining of a gel with ethidium bromide andvisualization under UV light. Alternatively, if the amplificationproducts are integrally labeled with radio- or fluorometrically-labelednucleotides, the amplification products can then be exposed to x-rayfilm or visualized under the appropriate stimulating spectra, followingseparation.

[0024] In one embodiment, visualization is achieved indirectly.Following separation of amplification products, a labeled, nucleic acidprobe is brought into contact with the amplified semenogelin sequence.The probe preferably is conjugated to a chromophore but may beradiolabeled. In another embodiment, the probe is conjugated to abinding partner, such as an antibody or biotin, where the other memberof the binding pair carries a detectable moiety (i.e., a label).

[0025] One example of the foregoing is described in U.S. Pat. No.5,279,721, which discloses an apparatus and method for the automatedelectrophoresis and transfer of nucleic acids. The apparatus permitselectrophoresis and blotting without external manipulation of the geland is ideally suited to carrying out methods according to the presentinvention.

[0026] It will be understood that the probes described above are limitedin as much as any nucleic-acid sequence can be used as long as thenucleic acid sequence is hybridizable to nucleic acids encodingsemenogelin or functional sequence analogs thereof. For example, anucleic acid of partial sequence can be used to quantify the expressionof a structurally related gene or the full-length genomic or cDNA clonefrom which it is derived.

[0027] Preferably, the hybridization is done under high stringencyconditions. By “high stringency conditions” is meant that the probespecifically hybridizes to a target sequence in an amount that isdetectably stronger than non-specific hybridization. High stringencyconditions, then, would be conditions which would distinguish apolynucleotide with an exact complementary sequence, or one containingonly a few scattered mismatches from a random sequence that happened tohave a few small regions (e.g., 3-10 bases) that matched the probe. Suchsmall regions of complementarity, are more easily melted than afull-length complement of 14-17 or more bases and high stringencyhybridization makes them easily distinguishable. Relatively highstringency conditions would include, for example, low salt and/or hightemperature conditions, such as provided by about 0.02-0.1 M NaCl or theequivalent, at temperatures of about 50-70° C. Such high stringencyconditions tolerate little, if any, mismatch between the probe and thetemplate or target strand, and are particularly suitable for detectingexpression of specific semenogelins. It is generally appreciated thatconditions can be rendered more stringent by the addition of increasingamounts of formamide.

[0028] When a semenogelin protein or polypeptide fragment thereof isassayed, various assays (i.e., immunobinding assays) are contemplated toeither detect or measure the quantity of semenogelin. For example, thesemenogelin can comprise the amino acid sequence of SEQ ID NO:1, SEQ IDNO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7,SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ IDNO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ IDNO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, or SEQ ID NO:27. Ifdesired, the semenogelin can comprise an amino acid sequence, which isspecific for SgI or SgII. For example, the semenogelin assayed for canbe SgI and can comprise the amino acid sequence of SEQ ID NO:2, SEQ IDNO:4, and SEQ ID NO:6. Alternatively, the semenogelin assayed for can beSgII and can comprise the amino acid sequence of SEQ ID NO:1, SEQ IDNO:3, and SEQ ID NO:5. In such embodiments, the semenogelin can beemployed to detect antibodies having reactivity therewith, or,alternatively, antibodies can be prepared and employed to detect thesemenogelin. The steps of various useful immunodetection assays havebeen described in Nakamura et al., Handbook of Experimental Immunology(4th Ed.), Wol. 1, Chapter 27, Blackwell Scientific Publ., Oxford(1987); Nakamura et al., Enzyme Immunoassays: Heterogenous andHomogenous Systems, Chapter 27 (1987) and include Western hybridization(i.e., Western blots), immunoaffinity purification, immunoaffinitydetection, enzyme-linked immunosorbent assay (e.g., an ELISA), andradioimmunoassay. A microarray also can be used to detect or measure thelevels of semenogelin.

[0029] In general, the immunobinding assays involve obtaining a testsample suspected of containing a protein, peptide or antibodycorresponding to a semenogelin, and contacting the sample with anantibody in accordance with the present invention, as the case may be,under conditions effective to allow the formation of immunocomplexes.Indeed, a mammal can be diagnosed with a cancer by either detecting SgI,SgII, or an antibody that recognizes semenogelin, such as when a femalemammal is being tested, or by quantifying the levels of SgI, SgII, or anantibody that recognizes semenogelin, such as when a male mammal isbeing tested.

[0030] Any suitable antibody can be used in conjunction with the presentinvention such that the antibody is specific for semenogelin, and inparticular, SgI and/or SgII. One known antibody which has suchspecificity is MHS-5. Indeed, MES-5 is a monoclonal antibody that canrecognize both SgI and SgII, as well as several proteolytic fragmentsthereof. Additionally, the antibody can recognize other antibodiespresent in a test sample that bind to semenogelin. Indeed, when such ananti-idiotypic antibody is used, it will typically recognize MHS-5.

[0031] The immunobinding assays for use in the present invention includemethods for detecting or quantifying the amount of semenogelin in asample, which methods require the detection or quantitation of anyimmune complexes formed during the binding process. Here, a test samplesuspected of containing a semenogelin would be obtained from a mammaland subsequently contacted with an antibody (e.g., MHS-5). The detectionor the quantification of the amount of immune complexes formed under thespecific conditions is then performed.

[0032] Contacting the biological sample with an antibody that recognizesa semenogelin under conditions effective and for a period of timesufficient to allow formation of immune complexes (primary immunecomplexes) is generally a matter of simply adding the antibody to thesample and incubating the mixture for a period of time long enough forthe antibodies to form immune complexes with, i.e., to bind to, anyantigens. After this time, the sample-antibody composition, such as atissue section, ELISA plate, dot blot or Western blot, will generally bewashed to remove any non-specifically bound antibody species, allowingonly those antibodies specifically bound within the primary immunecomplexes to be detected.

[0033] In general, the detection of immunocomplex formation iswell-known in the art and can be achieved through the application ofnumerous approaches. These methods are generally based upon thedetection of a label or marker, such as any radioactive, fluorescent,biological or enzymatic tags or labels of standard use in the art. U.S.Patents concerning the use of such labels include U.S. Pat. Nos.3,817,837, 3,850,752, 3,939,350, 3,996,345, 4,277,437, 4,275,149 and4,366,241. Of course, additional advantages can be realized by using asecondary binding ligand, such as a second antibody or a biotin/avidinligand binding arrangement, as is known in the art.

[0034] The antibody which is used in the context of the presentinvention can, itself, be linked to a detectable label, wherein onewould then simply detect this label, thereby allowing the presence of orthe amount of the primary immune complexes to be determined.

[0035] Alternatively, the first added component that becomes boundwithin the primary immune complexes can be detected by means of a secondbinding ligand that has binding affinity for the first antibody. Inthese cases, the second binding ligand is, itself, often an antibody,which can be termed a “secondary” antibody. The primary immune complexesare contacted with the labeled, secondary binding ligand, or antibody,under conditions effective and for a period of time sufficient to allowthe formation of secondary immune complexes. The secondary immunecomplexes are then washed to remove any non-specifically bound labeledsecondary antibodies or ligands, and the remaining label in thesecondary immune complexes is then detected.

[0036] Further methods include the detection of primary immune complexesby a two-step approach. A second binding ligand, such as an antibody,that has binding affinity for the first antibody is used to formsecondary immune complexes, as described above. After washing, thesecondary immune complexes are contacted with a third binding ligand orantibody that has binding affinity for the second antibody, again underconditions effective and for a period of time sufficient to allow theformation of immune complexes (tertiary immune complexes). The thirdligand or antibody is linked to a detectable label, allowing detectionof the tertiary immune complexes thus formed.

[0037] It will be understood that other diagnostic tests can be used inconjunction with the diagnostic tests described herein to enhancefurther the accuracy of diagnosing a mammal with a cancer. For example,a monoclonal antibody, such as the ones described in U.S. Pat. No.4,569,788 can be used effectively in diagnosing small-cell lung cancerover non small-cell lung cancer.

[0038] The present invention also provides a method of prognosticating acancer in a mammal, wherein the cancer is other than prostate cancer andsemenogelin is a marker for the cancer, which method comprises measuringthe level of semenogelin in a test sample obtained from the mammal,wherein the level of semenogelin in the test sample is indicative of theprognosis of the cancer in a mammal. The level of semenogelin in thetest sample can be measured by comparing the level of semenogelin inanother test sample obtained from the mammal over time in accordancewith the methods described above. An increase in semenogelin from onesample to the next is indicative of growth and/or metastasis of thecancer, whereas a decrease in semenogelin from one sample to the next isindicative of reduction of the cancer.

[0039] The invention also provides a method of assessing theeffectiveness of treatment of a cancer in a mammal, wherein the canceris other than prostate cancer and semenogelin is a marker for thecancer, which method comprises measuring the level of semenogelin in atest sample obtained from the mammal, wherein the level of semenogelinin the test sample is indicative of the effectiveness of the treatmentof the cancer in the mammal. The level of semenogelin in the test samplecan be measured by comparing the level of semenogelin in the test sampleto the level of semenogelin in another test sample obtained from themammal over time in accordance with the methods described above. Anincrease in semenogelin from one sample to the next is indicative of thetreatment being ineffective, whereas no change or a decrease insemenogelin from one sample to the next is indicative of the treatmentbeing effective.

[0040] Although semenogelin is known to be a secreted protein,semenogelins are expressed ectopically by cancer cell lines, inparticular SCLC cell lines derived from both male and female mammals.Furthermore, membrane association of semenogelin proteins is enhanced bythe addition of epidermal growth factor (EGF) when the cells areattached to thrombospondin-1 peptides. Thrombospondin-1 is a peptideknown to bind to integrins on a tumor cell surface. Integrins arecell-surface receptors responsible for the attachment of cells to theextracellular matrix. As a result, semenogelin associates with athrombospondin-1 peptide, which, in turn, associates with an integrin onthe surface of the tumor cell. This process is enhanced with theaddition of EGF. Accordingly, the present invention further provides amethod of inducing an immune response to a cancer in a mammal, whereinthe cancer is other than prostate cancer and semenogelin is a marker forthe cancer. The method comprises administering to the mammal acomposition comprising (a) an immune-response inducing effective amountof (i) a semenogelin protein or polypeptide fragment thereof, or (ii) anantibody or antigenically reactive fragment thereof that is specific fora semenogelin protein or polypeptide fragment thereof, or (b) arecombinant vector encoding and expressing an immune-response inducingeffective amount of (i) or (ii), whereupon an immune response to thecancer is induced. For example, the method of inducing an immuneresponse to the mammal can comprise administering to the mammal acomposition comprising (a) an imnmune-response inducing effective amountof (i) a polypeptide of SEQ D NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ DNO:4, SEQ ID NO:5, SEQ D NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9,SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14,SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19,SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24,SEQ ID NO:25, SEQ ID NO:26, or SEQ ID NO:27 or (ii) an antibody orantigenically reactive fragment thereof that is specific for apolypeptide of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ IDNO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ IDNO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ IDNO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ IDNO:25, SEQ ID NO:26, or SEQ ID NO:27 or (b) a recombinant vectorencoding and expressing an immune-response inducing effective amount of(i) or (ii), whereupon an immune response to the cancer is induced.

[0041] The present invention also provides a composition. Thecomposition comprises a pharmaceutically acceptable carrier and (a) animmune-response inducing effective amount of (i) a polypeptide of SEQ IDNO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6,SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:1, SEQ IDNO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ IDNO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ IDNO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, or SEQ IDNO:27 or (ii) an antibody or antigenically reactive fragment thereofthat is specific for a polypeptide of SEQ ID NO:1, SEQ ID NO:2, SEQ BDNO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8,SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:1, SEQ ID NO:12, SEQ ID NO:13, SEQID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ IDNO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ IDNO:24, SEQ ID NO:25, SEQ ID NO:26, or SEQ ID NO:27 or (b) a recombinantvector encoding and expressing an immune-response inducing effectiveamount of (i) or (ii).

[0042] The semenogelin protein or polypeptide fragment thereof of (i)and the antibody or antigenically reactive fragment thereof of (ii) canbe purified and isolated from a naturally occurring source or can besynthetically produced. Alternatively, the semenogelin protein orpolypeptide fragment thereof of (i) and the antibody or antigenicallyreactive fragment thereof of (ii) can be recombinantly produced. In thisrespect, the recombinant vector of (b) comprises a nucleic acid sequencewhich encodes and expresses an immune-response inducing effective amountof (i) or (ii). Methods of recombinant production and synthesis areknown in the art (see, e.g., Sambrook et al. (1989), supra) as aremethods of antibody production (see, e.g., Antibodies: A LaboratoryManual, Cold Spring Harbor Laboratory, (1988)). In accordance with thepresent invention, an antigenically reactive fragment of the antibody(e.g., Fab, Fc, etc.) can be obtained from the antibodies produced asdescribed above, by methods which include digestion with enzymes, suchas pepsin or papain, and/or cleavage of disulfide bonds by chemicalreduction.

[0043] If the semenogelin protein or polypeptide fragment of (i) and theantibody or antigenically reactive fragment thereof of (ii) arerecombinantly produced, any suitable recombinant vector can be used.Suitable vectors include, for example, plasmid vectors, retroviralvectors, adenoviral vectors, adeno-associated viral vectors, vacciniavirus, sindbis virus, cytomegalovirus, herpes simplex virus, defectivehepatitis B viruses, and any other vector or vector system known in theart.

[0044] The composition can comprise more than one active ingredient,such as more than one semenogelin of the present invention.Alternatively, or additionally, the composition can comprise anotherpharmaceutically active agent or drug.

[0045] The carrier can be any suitable carrier. Preferably, the carrieris a pharmaceutically acceptable carrier. With respect to compositions,the carrier can be any of those conventionally used and is limited onlyby chemico-physical considerations, such as solubility and lack ofreactivity with semenogelin, and by the route of administration. It willbe appreciated by one of skill in the art that, in addition to theabove-described composition, the compositions of the present inventivemethods can be formulated as inclusion complexes, such as cyclodextrininclusion complexes, or liposomes.

[0046] The pharmaceutically acceptable carriers described herein, forexample, vehicles, adjuvants, excipients, and diluents, are well-knownto those skilled in the art and are readily available to the public. Itis preferred that the pharmaceutically acceptable carrier be one whichis chemically inert to the semenogelin and one which has no detrimentalside effects or toxicity under the conditions of use.

[0047] The choice of carrier will be determined in part by theparticular semenogelin, as well as by the particular method used toadminister the composition. Accordingly, there are a variety of suitableformulations of the composition of the present invention. The followingformulations for oral, aerosol, parenteral, subcutaneous, intravenous,intramuscular, interperitoneal, rectal, and vaginal administration areexemplary and are in no way limiting.

[0048] The method of inducing an immune response to a cancer using acomposition of the present invention can be made more effective byadministering one or more other anticancer compounds along with one ormore other compositions of the present invention. These other anticancercompounds include, but are not limited to, all of the known anticancercompounds approved for marketing in the United States and those thatwill become approved in the future. See, for example, Table 1 and Table2 of Boyd, Current Therapy in Oncology, Section I. Introduction toCancer Therapy (J. E. Niederhuber, ed.), Chapter 2, by B. C. Decker,Inc., Philadelphia, 1993, pp. 11-22. More particularly, these otheranticancer compounds include doxorubicin, bleomycin, vincristine,vinblastine, VP-16, VW-26, cisplatin, carboplatin, procarbazine, andtaxol for solid tumors in general; alkylating agents, such as BCNU,CCNU, methyl-CCNU and DTIC, for brain or kidney cancers; andantimetabolites such as 5-FU and methotrexate for colon cancer.

[0049] One skilled in the art will appreciate that suitable methods ofadministering a composition of the invention to a mammal, in particulara human, are available, and, although more than one route can be used toadminister a particular compound, a particular route can provide a moreimmediate and more effective reaction than another route. Accordingly,the herein-described methods are exemplary and are in no way limiting.

[0050] The dose administered to a mammal, in particular a human, shouldbe sufficient to induce an immune response to the cancer. One skilled inthe art will recognize that dosage will depend upon a variety of factorsincluding the strength of the particular composition employed, as wellas the age, species, condition, and body weight of the mammal. The sizeof the dose will also be determined by the route, timing, and frequencyof administration as well as the existence, nature, and extent of anyadverse side-effects that might accompany the administration of aparticular composition and the desired physiological effect.

[0051] Suitable doses and dosage regimens can be determined byconventional range-finding techniques known to those of ordinary skillin the art. Generally, a composition is initially administered insmaller dosages, which are less than the optimum dose of thecomposition. Thereafter, the dosage is increased by small incrementsuntil the optimum effect under the circumstances is reached. The presentinventive method will typically involve the administration of about0.1-100 mg of one or more of the compositions described above per kgbody weight.

EXAMPLES

[0052] The following examples further illustrate the invention but, ofcourse, should not be construed as in any way limiting its scope.

[0053] A number of assays are used in the examples below, includingisoelectric focusing, 2-dimensional gel electrophoresis,SDS-polyacrylamide gel electrophoresis, Western blotting, massspectrometry, and liquid chromatography. The first four are described inAlberts et al., Molecular Biology of the Cell, 3^(rd) Ed., GarlandPublishing, Inc., New York, N.Y. (1994), mass spectrometry is describedin Skoog et al., Principles of Instrumental Analysis, Holt, Rinehart andWinston, Inc. (1971), and liquid chromatography is described inMohrig-Neckers, Laboratory Experiments in Organic Chemistry, 2^(nd) Ed.Litton Educational Publishing, Inc., New York, N.Y. (1973).

Example 1

[0054] This example demonstrates that semenogelin is expressedectopically by SCLC and melanoma cell lines.

[0055] A synthetic peptide containing the thrombospondin-1 sequence,FQGVLQNVRFVF (peptide 678; SEQ ID NO:28), that binds to the αβ1 integrinwas prepared as described in Krutzsch et al., J. Biol. Chem.:24080-24086 (1999). Semenogelin was purified from seminal plasma asdescribed in Malm, Eur. J. Biochem. 238: 48-53 (1996), and recombinantEGF was obtained from R & D Systems (Minneapolis, Minn.) for use in thefollowing assays.

[0056] Two different types of cancer cell lines were tested forsemenogelin. Specifically, lung cancer (OH-1, NCI-N592, NCI-N417,NCI-H378, NCI-H570, NCI-H727, NCI-H157, NCI-H520, NCI-A549) and melanoma(C32, A2058) cell lines were grown in RPMI 1640 medium containing 10%fetal calf serum (15% FCS for OH-1 cells) until log phase.

[0057] The lung cancer and melanoma cell lines' were then dissociated byreplacing the growth medium with 2.5 mM EDTA in phosphate-bufferedsaline and incubating at 37° C. for 10 minutes. Cells were thentriturated and collected by centrifugation, suspended in M199 medium,and plated on thrombospondin-1 peptide coated plates (Falcon 1029). Celladhesion to thrombospondin-1 peptide 678 (10 p was assessed with andwithout EGF (5 ng/ml). Cells were incubated for 2 hours at 37° C. andthen aspirated and washed three times with Dulbecco's PBS.Cytoskeleton-associated adhesion complexes were isolated by detergentextraction of the cells using CSK buffer (0.5% Triton-X-100, 50 mM NaCl,300 mM sucrose, 3 mM MgCl2, 20 μg/ml aprotinin, 1 μg/ml leupeptin, 1μg/ml pepstatin, 1 mM phenylmethanesulfonyl fluoride, 10 mMpiperazine-N,N′-bis-(2-ethanesulfonic acid), pH 6.8) for 1 minutefollowed by sonication for 30 seconds to remove cell bodies. Thecell-surface adhesion complexes, which remained attached to the peptidesubstrates, were then extracted using 0.5 mL of IX RIPA buffer (50 mMTris pH 7.2, 1% Triton-X-100, 1% deoxycholate, 0.1% SDS, 150 mM NaCl, 1mM phenylmethanesulfonyl fluoride). The plates were scraped, therecovered extracts were centrifuged, and the supernatant fractions werecollected.

[0058] Proteins extracted from the thrombospondin-1 peptide-associatedadhesion complexes were mixed with an equal volume of rehydration buffer(8 M urea, 1% IPG, 2% CHAPS in H₂O) in a strip holder with a gel strip,pH 3-10 (Amersham Pharmacia Biotech, Piscataway, N.J.) and incubatedovernight at 20° C. The strip holders and hydrated strips were placed inan IPGphor unit and subjected to isoelectric focusing for 4 hours. Thegels were then equilibrated with 100 mg dithiothreitol in 10 ml H₂O for1 hour, followed by the addition of 100 mg of iodoacetamide and shakingfor 15 minutes. The equilibrated gel strip was applied to a sodiumdodecyl sulfate/10% polyacrylamide gel and subjected to electrophoresis(i.e., 2-D gel electrophoresis) on a Hoefer TE Transphor for 15 minutesat 20 mA, then 5 hours at 40 mA. The gels were removed and stained withCoomassie blue stain for 1 hour and destained (10% acetic acid +35%methanol in H₂O) overnight.

[0059] Analysis of adhesion complexes isolated from both lung cancer andmelanoma cells by 2-dimensional gel electrophoresis showed that twoproteins with molecular weights of 25-58 kDa and a pI of 9 were inducedto associate with the adhesion complexes by the addition of EGF. The twoprotein spots were isolated and sequenced by mass spectrometry and foundto contain multiple peptides homologous to portions of SgI and SgII.

[0060] As indicated by these results, semenogelin is expressedectopically by SCLC and melanoma cell lines. Moreover, the membraneassociation of the semenogelin proteins to a cancer cell was enhanced bythe addition of EGF when the cancer cell(s) was attached tothrombospondin-1 peptides via an αβB1 integrin.

Example 2

[0061] This example also demonstrates that semenogelin is expressed inlung cancer and melanoma cell lines.

[0062] Lung cancer and melanoma cell lines were harvested and extractedas described above. Extracted proteins, after clarifying bycentrifugation, were separated by SDS-polyacrylamide gel electrophoresisin Mini Protean II System (BioRad-Hercules, Calif.) for 55 minutes at170V. Following electrophoresis, proteins were transferred ontopolyvinylidine difluoride membrane in transfer buffer (20% methanol in1×Tris/glycine, BioRad) at 70V for 2.5 hours and washed three times with1×Tris/glycine buffer. The membrane was blocked overnight in Dulbecco'sPBS containing 1% bovine serum albumin and 0.1% Tween-20. The blot wasthen incubated with biotinylated MHS-5 antibody (Humagen FertilityBiagnostics, Charlottesville, Va.) at 1:1000 for 2 hours at 37° C.followed by four washes with Dulbecco's PBS containing 0.1% Tween. Themembrane was then incubated with streptavidin-horseradish peroxidase(Amersham Pharmacia Biotech) diluted 1:20,000 for 1 hour. Surfaceproteins were visualized with an chemiluminescent detection kit(Amersham Pharmacia Biotech).

[0063] Additionally, unlabeled MHS-5 antibody was pre-bound to Protein Aagarose in Dulbecco's PBS containing 1% BSA and 0.1% Tween-20 for 2hours at 4° C. Beads containing the bound antibody were then incubatedwith 500 μL of extracted proteins overnight at 4° C. The beads werewashed three times with Tris-buffered saline, then eluted by heatingwith sample buffer (50 mM Tris, 6 M urea, 30% glycerol, 2% SDS, andbromophenol blue) at 95° C. for 5 minutes, and separated and transferredas described previously. The blot was then incubated with biotinylatedMHS-5 antibody, streptavidin-horseradish peroxidase, and visualized bychemiluminescence.

[0064] Proteins of interest contained in 1- or 2-dimensional gels wereanalyzed by liquid chromatography/mass spectrometry (LC/MS) to determinetheir identity. The protein spot detected in the stained gel was excisedand placed in a microfuge tube. The gel piece was washed withmethanol/ammonium bicarbonate buffer, dried in vacuo, then treated withtrypsin overnight. The resulting peptides were extracted, separated, andanalyzed on a Finnigan LCQ LC/MS system. The resulting run files werefirst analyzed using Sequest database searching software. If noidentification resulted, then further database searching and/or de novosequencing was carried out.

[0065] Mass spectrometric analyses of tryptic peptides from these twofragments yielded peptides that covered 42.6% of the SgI proteinsequence and 38.5% of the SgII protein sequence. Several peptidesdetected were identical in both proteins, as expected from their knownsequence homologies. These peptides are described in Table 1. However,several of the ion peaks were assigned to peptides that could beassigned specifically to semenogelin I or II. These resultingpolypeptides are described in Table 2. TABLE 1 Position in Complete SEQID Amino Acid NO: Peptide Sequence Sequence 7 ISYQSSSTEER 406-416 8IPSQAQEYGHK 392-402 9 GQHYFGQK 45-52 10 SQNQVTIPSQDQEHGHK 446-462 11KSQQYDLNALHK 81-92 12 SQIQTPNPNQDQWSGQNAK 506-524 13 GSISIQTEEKIHGK432-445 14 SQQYDLNALHK 82-92 15 GHFHMIVIHHK 121-131 16 LWVHGLSK 166-17317 DVSQSSISFQIEK 488-500 18 GSFSIQHTYHVDINDHDWTR 61-80 19TQGGSQSSYVLQTEELVVNK 186-205 20 HLAQHLNNDR 448-457 21 GISSQYSNTEER154-165 22 LHYGENGVQK 358-367 23 DVSQSSIYSQTEEK 308-321 24QITIPSQEQEHSQK 329-342 25 LPSEFSQFPHGQK 32-44 26 HQHGSHGGLDIVIIEQEDDSDR426-447 27 DIFSTQDELLVYNK 252-265

[0066] TABLE 2 SEQ Position in Complete ID Peptide Amino Acid NO:Sequence Semenogelin Sequence 1 QHLGGSQQLLNYK SgII  98-110 2HLGGSQQLLHNK SgI   98-110 3 GHYQNVVDVR SgII 218-227 4 GHYQNVVEVR SgI 218-227 5 QDLLSHEQK SgII 535-543 6 EQDLLSHEQK SgI  414-423

[0067] Nearly all SCLC lines screened by Western blotting demonstratedthe presence of a doublet between 70-80 kDa corresponding to SgII. Onlythe classic SCLC cell line in OH-1, which grows as tight aggregates invitro, was positive for SgI. SgII was detected in all of the SCLC andsome non-SCLC lung cancer cell lines, including some squamous cellcarcinomas. SgII expression was also detected in two melanoma cell lines(A2058 and C32 cell lines).

[0068] Furthermore, SgII was found in cell lines derived from SCLCpatients of either gender. Expression of SgII in the female lines H378and N417 further supports specific ectopic production of thesemenogelins by the SCLC cells.

[0069] SgII was also secreted into the medium by OH-1 SCLC cells.Measurement of secreted SgII with MHS-5, therefore, can provide aquantitative method to assess SCLC tumor burden.

[0070] These results suggest that both semenogelins were cleavedsomewhere in the middle of the protein to generate the observedfragments set forth in Tables 1 and 2 and evidence that semenogelin isexpressed in both lung cancer and melanoma cell lines.

Example 3

[0071] This example demonstrates that semenogelin can be detected intumor tissue samples taken from patients diagnosed with SCLC.

[0072] Formalin-fixed paraffin-embedded representative tissue specimenswere taken from 13 specimens and immunostained with monoclonal antibodyagainst MHS-5 (dilution 1/100). Staining was performed with the EnVisionsystem (DAKO). Endogenous peroxidase activity was quenched by treatmentwith 5% hydrogen peroxide in methanol for 30 minutes at roomtemperature. Antigen retrieval using Target Retrieval pH 7.0 (DAKO) andmicrowave treatment for 20 minutes in an 800-watt microwave open wasperformed. A blocking step with protein block serum free (DAKO) wasused. The primary antibody was then applied for 120 minutes at roomtemperature. The sections were rinsed with washing buffer (DulbeccosPBS+0.1% Tween) at room temperature and incubated with EnVision systemreagents for 30 minutes at room temperature. The slides were rinsed withwashing buffer, and treated with a solution containing 0.05%diaminobenzidine hydrochloride and 0.1% hydrogen peroxide in 0.05 mol/lTRIS-buffered saline, pH 7.4, at room temperature for 5 minutes. Afterrinsing in distilled water for 3 minutes, the slides were counterstainedwith modified Harris hematoxylin, dehydrated, and mounted. Negativecontrol sections were treated in an identical fashion except for lack ofprimary antibody. An appropriate positive control (human seminalvesicle) was run concurrently. Of the 13 SCLC specimens analyzed, 12were found to be diffusely positive with homogenous, widespread antibodylabeling throughout the cytoplasm of the tumor cells and surroundingmatrix. These results indicate that semenogelin is ectopically expressedin SCLC cells taken directly from an individual and, thus, furtherevidence that semenogelin is an effective marker in diagnosing SCLC.

[0073] All of the references cited herein, including patents, patentapplications, and publications, are hereby incorporated in theirentireties by reference.

[0074] While this invention has been described with an emphasis uponpreferred embodiments, variations of the preferred embodiments can beused, and it is intended that the invention can be practiced otherwisethan as specifically described herein. Accordingly, this inventionincludes all modifications encompassed within the spirit and scope ofthe invention as defined by the claims.

1 28 1 13 PRT Homo sapiens 1 Gln His Leu Gly Gly Ser Gln Gln Leu Leu AsnTyr Lys 1 5 10 2 12 PRT Homo sapiens 2 His Leu Gly Gly Ser Gln Gln LeuLeu His Asn Lys 1 5 10 3 10 PRT Homo sapiens 3 Gly His Tyr Gln Asn ValVal Asp Val Arg 1 5 10 4 10 PRT Homo sapiens 4 Gly His Tyr Gln Asn ValVal Glu Val Arg 1 5 10 5 9 PRT Homo sapiens 5 Gln Asp Leu Leu Ser HisGlu Gln Lys 1 5 6 10 PRT Homo sapiens 6 Glu Gln Asp Leu Leu Ser His GluGln Lys 1 5 10 7 11 PRT Homo sapiens 7 Ile Ser Tyr Gln Ser Ser Ser ThrGlu Glu Arg 1 5 10 8 11 PRT Homo sapiens 8 Ile Pro Ser Gln Ala Gln GluTyr Gly His Lys 1 5 10 9 8 PRT Homo sapiens 9 Gly Gln His Tyr Phe GlyGln Lys 1 5 10 17 PRT Homo sapiens 10 Ser Gln Asn Gln Val Thr Ile ProSer Gln Asp Gln Glu His Gly His 1 5 10 15 Lys 11 12 PRT Homo sapiens 11Lys Ser Gln Gln Tyr Asp Leu Asn Ala Leu His Lys 1 5 10 12 19 PRT Homosapiens 12 Ser Gln Ile Gln Thr Pro Asn Pro Asn Gln Asp Gln Trp Ser GlyGln 1 5 10 15 Asn Ala Lys 13 14 PRT Homo sapiens 13 Gly Ser Ile Ser IleGln Thr Glu Glu Lys Ile His Gly Lys 1 5 10 14 11 PRT Homo sapiens 14 SerGln Gln Tyr Asp Leu Asn Ala Leu His Lys 1 5 10 15 11 PRT Homo sapiens 15Gly His Phe His Met Ile Val Ile His His Lys 1 5 10 16 8 PRT Homo sapiens16 Leu Trp Val His Gly Leu Ser Lys 1 5 17 13 PRT Homo sapiens 17 Asp ValSer Gln Ser Ser Ile Ser Phe Gln Ile Glu Lys 1 5 10 18 20 PRT Homosapiens 18 Gly Ser Phe Ser Ile Gln His Thr Tyr His Val Asp Ile Asn AspHis 1 5 10 15 Asp Trp Thr Arg 20 19 20 PRT Homo sapiens 19 Thr Gln GlyGly Ser Gln Ser Ser Tyr Val Leu Gln Thr Glu Glu Leu 1 5 10 15 Val ValAsn Lys 20 20 10 PRT Homo sapiens 20 His Leu Ala Gln His Leu Asn Asn AspArg 1 5 10 21 12 PRT Homo sapiens 21 Gly Ile Ser Ser Gln Tyr Ser Asn ThrGlu Glu Arg 1 5 10 22 10 PRT Homo sapiens 22 Leu His Tyr Gly Glu Asn GlyVal Gln Lys 1 5 10 23 14 PRT Homo sapiens 23 Asp Val Ser Gln Ser Ser IleTyr Ser Gln Thr Glu Glu Lys 1 5 10 24 14 PRT Homo sapiens 24 Gln Ile ThrIle Pro Ser Gln Glu Gln Glu His Ser Gln Lys 1 5 10 25 13 PRT Homosapiens 25 Leu Pro Ser Glu Phe Ser Gln Phe Pro His Gly Gln Lys 1 5 10 2622 PRT Homo sapiens 26 His Gln His Gly Ser His Gly Gly Leu Asp Ile ValIle Ile Glu Gln 1 5 10 15 Glu Asp Asp Ser Asp Arg 20 27 14 PRT Homosapiens 27 Asp Ile Phe Ser Thr Gln Asp Glu Leu Leu Val Tyr Asn Lys 1 510 28 12 PRT Artificial Synthetic peptide 28 Phe Gln Gly Val Leu Gln AsnVal Arg Phe Val Phe 1 5 10

1. A method of diagnosing cancer in a male mammal, wherein the cancer isother than prostate cancer, which method comprises: (a) obtaining a testsample from the male mammal, and (b) assaying the test sample for anincreased level of semenogelin, wherein the increased level ofsemenogelin in the test sample is diagnostic for the cancer.
 2. Themethod of claim 1, wherein the test sample is assayed for an increasedlevel of semenogelin in (b) by comparing the level of semenogelin in thetest sample to the level of semenogelin in a control sample obtainedfrom one or more cancer-free male mammals of the same species, whereinan increase in the level of semenogelin in the test sample as comparedto the control sample is diagnostic for the cancer.
 3. A method ofdiagnosing cancer in a female mammal, which method comprises: (a)obtaining a test sample from the female mammal, and (b) assaying thetest sample for the presence of semenogelin, wherein the presence ofsemenogelin in the test sample is diagnostic for the cancer.
 4. Themethod of claim 1, wherein the semenogelin comprises the amino acidsequence of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ IDNO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10,SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15,SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20,SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25,SEQ ID NO:26, or SEQ ID NO:27.
 5. The method of claim 1, wherein thesemenogelin is semenogelin I.
 6. The method of claim 5, wherein thesemenogelin I comprises the amino acid sequence of SEQ ID NO:2, SEQ IDNO:4 or SEQ ID NO:6.
 7. The method of claim 1, wherein the semenogelinis semenogelin II.
 8. The method of claim 7, wherein the semenogelin IIcomprises the amino acid sequence of SEQ ID NO:1, SEQ ID NO:3 or SEQ IDNO:5.
 9. The method of claim 1, wherein the test sample is serum ortissue.
 10. (Canceled)
 11. The method of claim 9, wherein the tissue istumor tissue.
 12. The method of claim 1, wherein (b) comprises assayingfor RNA encoding semenogelin.
 13. (Canceled)
 14. The method of claim 1,wherein (b) comprises-assaying for semenogelin protein or a polypeptidefragment thereof.
 15. (Canceled)
 16. The method of claim 1, wherein thecancer is lung cancer and the method further comprises (c) testing forthe presence of small-cell lung cancer.
 17. A method of prognosticatinga cancer in a mammal, wherein the cancer is other than prostate cancerand semenogelin is a marker for the cancer, which method comprisesmeasuring the level of semenogelin in a test sample obtained from themammal, wherein the level of semenogelin in the test sample isindicative of the prognosis of the cancer in the mammal.
 18. The methodof claim 17, wherein the level of semenogelin in the test sample ismeasured by comparing the level of semenogelin in the test sample to thelevel of semenogelin in another test sample obtained from the mammalover time.
 19. A method of assessing the effectiveness of treatment of acancer in a mammal, wherein the cancer is other than prostate cancer andsemenogelin is a marker for the cancer, which method comprises measuringthe level of semenogelin in a test sample obtained from the mammal,wherein the level of semenogelin in the test sample is indicative of theeffectiveness of the treatment of the cancer in the mammal.
 20. Themethod of claim 19, wherein the level of semenogelin in the test sampleis measured by comparing the level of semenogelin in the test sample tothe level of semenogelin in another test sample obtained from the mammalover time.
 21. A method of inducing an immune response to a cancer in amammal, wherein the cancer is other than prostate cancer and semenogelinis a marker for the cancer, which method comprises administering to themammal a composition comprising (a) an immune-response inducingeffective amount of (i) a semenogelin protein or polypeptide fragmentthereof or (ii) an antibody or antigenically reactive fragment thereofthat is specific for a semenogelin protein or polypeptide fragmentthereof or (b) a recombinant vector encoding and expressing animmune-response inducing effective amount of (i) or (ii), whereupon animmune response to the cancer is induced.
 22. The method of claim 21,wherein the composition comprises (a) an immune-response inducingeffective amount of (i) a polypeptide of SEQ ID NO:1, SEQ ID NO:2, SEQID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ IDNO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ IDNO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ IDNO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ IDNO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, or SEQ ID NO:27 or (ii)an antibody or antigenically reactive fragment thereof that is specificfor a polypeptide of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4,SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ IDNO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ IDNO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ IDNO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ IDNO:25, SEQ ID NO:26, or SEQ ID NO:27 or (b) a recombinant vectorencoding and expressing an immune-response inducing effective amount of(i) or (ii), whereupon an immune response to the cancer is induced. 23.The method of claim 1, wherein the cancer is of epithelial origin. 24.The method of claim 23, wherein the cancer is lung cancer.
 25. Themethod of claim 24, wherein the lung cancer is small-cell lung cancer.26. The method of claim 1, wherein the cancer is melanoma.
 27. Acomposition comprising a pharmaceutically acceptable carrier and (a) animmune-response inducing effective amount of (i) a polypeptide of SEQ IDNO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6,SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ IDNO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ IDNO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, or SEQ IDNO:27 or (ii) an antibody or antigenically reactive fragment thereofthat is specific for a polypeptide of SEQ ID NO:1, SEQ ID NO:2, SEQ IDNO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8,SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ IDNO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ IDNO:24, SEQ ID NO:25, SEQ ID NO:26, or SEQ ID NO:27 or (b) a recombinantvector encoding and expressing an immune-response effective amount of(i) or (ii).
 28. The method of claim 3, wherein the semenogelincomprises the amino acid sequence of SEQ ID NO:1, SEQ ID NO:2, SEQ IDNO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8,SEQ ID NO:9, SEQ ID NO: 10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13,SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18,SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23,SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, or SEQ ID NO:27.
 29. Themethod of claim 3, wherein the semenogelin is semenogelin I.
 30. Themethod of claim 29, wherein the semenogelin I comprises the amino acidsequence of SEQ ID NO:2, SEQ ID NO:4 or SEQ ID NO:6.
 31. The method ofclaim 3, wherein the semenogelin is semenogelin II.
 32. The method ofclaim 31, wherein the semenogelin II comprises the amino acid sequenceof SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:
 5. 33. The method of claim 3,wherein the test sample is serum or tissue.
 34. The method of claim 33,wherein the tissue is tumor tissue.
 35. The method of claim 3, wherein(b) comprises assaying for RNA encoding semenogelin.
 36. The method ofclaim 3, wherein (b) comprises assaying for semenogelin protein or apolypeptide fragment thereof.
 37. The method of claim 3, wherein thecancer is lung cancer and the method further comprises (c) testing forthe presence of small-cell lung cancer.
 38. The method of claim 3,wherein the cancer is of epithelial origin.
 39. The method of claim 38,wherein the cancer is lung cancer.
 40. The method of claim 39, whereinthe lung cancer is small-cell lung cancer.
 41. The method of claim 3,wherein the cancer is melanoma.
 42. The method of claim 17, wherein thecancer is of epithelial origin.
 43. The method of claim 42, wherein thecancer is lung cancer.
 44. The method of claim 43, wherein the lungcancer is small-cell lung cancer.
 45. The method of claim 17, whereinthe cancer is melanoma.
 46. The method of claim 19, wherein the canceris of epithelial origin.
 47. The method of claim 46, wherein the canceris lung cancer.
 48. The method of claim 47, wherein the lung cancer issmall-cell lung cancer.
 49. The method of claim 19, wherein the canceris melanoma.
 50. The method of claim 21, wherein the cancer is ofepithelial origin.
 51. The method of claim 50, wherein the cancer islung cancer.
 52. The method of claim 51, wherein the lung cancer issmall-cell lung cancer.
 53. The method of claim 21, wherein the canceris melanoma.